There is a significant and growing need for the ex vivo creation of mammalian tissues for the augmentation or replacement of damaged tissues and organs. For example, cartilage cells produced in vitro may be useful for repairing cartilage that has been damaged in a knee injury or deteriorated by osteoarthritis.
The successful development of tissues created ex vivo depends on several factors. Such factors may include an adequate cell source that can be grown and differentiated into the desired tissue, a template that will promote cell adhesion and induce the deposition of extracellular matrix proteins, and a growth environment that will foster cell communication.
Different cells must possess different properties for proper functioning in the human body. For example, articular cartilage, or the cartilage that lines bones in joints, is firm and flexible connective tissue that is specialized to absorb and resist compression. Articular cartilage is protected by a nutritive and lubricating medium known as the synovial fluid of the joint. Cartilage is composed of chondrocyte cells, which occupy 10-20% of its volume, and an extracellular material that contains up to 80% water by weight.
The chondrocytes are enclosed within small cavities, called lacunae, generally in groups of 2, 4, or 6 cells as a result of mitosis and restricted cellular movement. The extracellular material consists primarily of large hydrated proteoglycan aggregates entrapped within a matrix of collagen fibrils. The matrix is predominantly made of type II collagen which forms a meshwork of high tensile-strength fibrils. The entrapped proteoglycans (also called mucopolysaccharides) are composed of a core protein that forms a backbone to which many glycosaminoglycan (GAG) chains are covalently attached. The GAGs are high negatively charged molecules that encourage the binding of water and the generation of a large osmotic swelling pressure.
The mechanical behavior of articular cartilage is similar to that of a sponge. During rest, for example when a person is sitting or lying down, the osmotic pressure generated by the proteoglycan aggregates fills the tissue with water up to its maximum capacity. This swelling pressure is contained only by the resilient collagen meshwork. Under load, such as when the person is standing up or walking, the weight of the body compresses the cartilage, squeezing water out until the osmotic pressure generated by the polyglycan produces a swelling force (due to the bound water) equal to the compressive force across the joint. When the load is removed, the cartilage slowly swells back to its full extent.
In order for the tissue created in vitro to function properly, it must have the same properties as the native tissue, or tissue found naturally in the human body. For example, experimental evidence has shown that the application of mechanical stimuli to engineered chondrocytic constructs that emulate the forces applied to articular cartilage leads to the production of a more functional artificially-produced tissue. In other words, tissue developed using the application of mechanical loading is more similar in its content and mechanical properties to physiologic cartilage than tissue produced without mechanical loading.
The main types of mechanical loads that have been investigated include (1) hydrostatic pressure, (2) direct mechanical compression, (3) high and/or low shear forces, (4) forced perfusion, and (5) ultrasonic compression or shear wave induction. Each of these involves the application of actual mechanical stimulation to the cells growing in vitro. A persistent drawback of each of these types of mechanical stimulation, however, is the increased complexity of the system necessary to produce the mechanical load stimulus. This complexity is magnified when production systems are scaled-up for producing large quantities of tissue.
Thus, there is a need for a bioreactor and method for producing and maintaining cells that are similar in content and possess similar mechanical properties to physiologic cells on a large scale with reduced complexity.